1. BACTERIAL IDENTIFICATION METHODS CHAPTER 3

2. Content • purification of cultures • morphological and pureculture studies • biochemical tests

3. Purification of cultures • The only way to purified culture are by getting a pure culture. • Have a control procedure in collecting specimen, preparing media, microbiological tecniques, staining and reagents and equipment used.

4. Collecting specimen • Sterile procedure using a correct media for transportation and stock. • Main source for bacteria should be microbial food. Which are?? • The transport mediaused to preserve and ensure the viability of bacteria during the transportation period • Besides also provide a particular temperature for the viability of bacteria • Crucial for cerebrospinal fluid, blood culture and fecal specimens, etc.

5. Preparation of media • The media should be packed well to prevent from leakage and breaks, protected from moisture and sunlight and excessive heat • The expiry date should be noted and the instruction of storage should be followed • The mix bacterial colonies should be sub cultured until the culture are purified • the bacterial colony characteristic should only derive from a single colony

6. Pure colony

7. Morphological and pure culture studies

8. In nutrient agar

10. In MacConkey agar

11. In MacConkey agar

12. In Blood agar

13. 3 types of hemolysis

14. Biochemical tests • Catalase test • Oxidase test • Coagulase test • Sugar fermentation test • MRVP test • Indole test • Citrate test • Motility test • H2S test

15. Catalase test • Produce bubble just after attaching the bacteria to the reagent • To differentiate staphylococci and streptococci

16. Oxidase test • Have 2 methods:Filter paper/Sterile swab • To help identify Vibrio, Neisseria, Pasteurellaand Pseudomonas sp. • Oxidase enzymes oxydizephenylenediamine. • Deep purple colour on reagent paper

17. Oxidase test

18. Coagulase test • To identify S.aureus • The enzyme coagulase clots plasma • Tube : fibrin clot • Slide: clumping of bacterial cells

19. Sugar fermentation test • Glucose test • Maltose test • Sucrose test • Lactose test • Some will appear with gas production

20. Voges-Proskauer test • To differentiate enterobacteria • Organism ferments glucose with acetoin production. Acetoin is oxidised to diacetylwhich reacts with creatine • Brick red colour develop slowly

21. Methyl Red test • To differentiate enterobacteria • Org. ferment glucose, producing sufficient acidity in buffered medium to give a colour change of indicator • Brick red medium

22. Indole test • Using Kovac reagent. • To differentiate Gram negative rods, especially E.coli • Tryptophan is broken down wt the release of indole which reacts wt dimethylaminobenzaldehyde • Reddening of strip or medium

23. Indole test using other reagent

24. Citrate test • To differentiate enterobacteria from other bacteria • Org uses citrate as its only source of carbon, producing an alkaline reaction wt colour change indicator • Blue and turbid medium

25. Motility test

26. The end Please do revision for your 1st test..next week!!